Two thermophilic bacteria were isolated from hot spring located near the city Chakwal in Pakistan. The cells of both strains were rod-shaped, stained Gram positive and formed endospores. The isolates were able to utilize sugars like fructose, Maltose, mannose, xylose, sacchorose, sorbitol, mannitol, Arabinose and polysaccharides like gelatin and starch. Phylogenetic analysis based on 16S rRNA gene sequences revealed that both isolates belong to the same specie Anoxybacillus rupiensis with 99% similarity under Accession number KF254911 (JF82) and KC849452 (JF83). The DNA G + C contents were 56.854% and 56.754% for A. rupiensis JF82 and A. rupiensis JF83, respectively. The isolates grew over a wide range of temperatures (40-70°C) and pH (4-9). The isolates exhibited growth with variety of nitrogen sources. Both thermophilic strains were grown on agro-dairy wastes to attain their cheap production by utilizing the wastes nutrients. Protein production was determined using 1% (w/v) of various agro-dairy wastes in production medium both with and without nutrients at pH 7.0 for 48 h at their respective suitable temperatures. They yielded enough thermostable protein suggesting their potential in production of various enzymes and proteins in unconventional and economical substrates suitable for various industrial uses.